For successful gene transfer to primitive hematopoietic cells several requirements need to be achieved. These include identification of the desired target cell population, identification of the appropriate vector to be used, and achieving desired levels of gene expression. To date, successful gene transfer in human subjects remain problematic. To address these problems as well as important safety issues, studies in non-human primates are being undertaken to optimize gene transfer to nonhuman primate hematopoietic cells prior to human clinical studies. Vectors that have been evaluated include self-inactivating (SIN) retroviral vectors, mst recently lentiviral vectors constructed to optimall transduce rhesus CD34+ cells. These vectors have been constructed to express reporter genes, such as the enhanced green fluorescent protein (EGFP), or therapeutic genes, such as hemoglobin. Transduction conditions employed the RGD-containing fibronectin fragment, RetroNectin (CH-296) and a variety of recombinant hematopoietic growth factors, such as stem cell factor (SCF), interleukin-6, megakaryocyte growth and differentiation factor (MGDF or thrombopoietin) and the human Flt-3 (fms-like tyrosine kinase) ligand in either serum containing or serum free media. Viral vectors evaluated include retroviral vectors, such as third generation chimeric human immunodeficiency virus type-1 (HIV-1)-based lentiviral vectors. Our efforts over the past year have resulted in publications evaluating the use of vectors in tracking lineage contributions of over time and the identification of genetic factors important in efficient gene trasduction of CD34+ cells. Efforts continue to be made to improve the level of gene marking, targeting gene expression to specific cell types, such as red blood cells, evaluate immune recostitution following transplant and the contribution of gentically marked cells to the recovery, and to derive stem cells from other tissues besides BM and cytokine mobilized PB, such as adult mesenchymal stem/progenitor cells and induced pluripotential stem cells and evaluate their safety in this in vivo model sytem. Recent success in developing induced pluripotential stem cells (iPS cells) as well as mesenchymal stromal cells (MSC) have been made this past year. Attempts are also being made to improve methodology and the technology behind stem cell mobilization and collection. Alternative approaches are being evaluated, for example, in both the instrumentation and methodology of leukapheresis procedures in small subjects. Despite continued improvements in methodology, questions remain. How can consistent high levels of expression be obtained using therapeutic genes? This is being evaluated by modifying globin expression in red blood cells. Can other stem cells either derived from bone marrow or other easily accessible tissues be targeted to assist in either the contribution or repair of other organs? This is being evaluated through the generation of iPS cells and MSC. How best to evaluate stem cells and their progeny therapeutically? Future studies are aimed to evaluate therapeutic vectors, improve hematopoietic stem cell recovery and transduction efficiency, further delineate the nature and clonality of populations contributing to the reconstitution using genetic tracking methodologies, and to isolate or induce and characterize primitive cell populations which may contribute to organogenesis or the repair of damaged tissues.